1. Field of the Invention
The present invention relates to a new therapeutic use of uridine in neuron degenerative diseases resulting from pathological ageing or from functional losses due to various causes, for example peripheral neuropathies, lateral amyotrophic sclerosis and Alzheimer's disease. This invention follows the evidence of the effects of uridine administering on various types of cells cultured.
According to the present invention in fact, uridine can act as a growth promoter when added to cell cultures, producing different effects due to the dose-levels (high or rather low) and the type of cells used as a target, and in particular when administered to neuronal and glial cell lines, uridine has the same biological effects as Nerve Growth Factor.
2. Description of the Prior Art
Growth factors constitute a large family of proteins fundamentally devoted to reproduction, differentiation, maturation and survival of cells. In the last few years, the tremendous developments in protein production through genetic engineering and biological techniques led to important discoveries on the physio-pathologic roles in mammalian bodies of several growth factors: as hyper- or hypo-production of growth factors by nearby or circulating cells has been linked to a large number of unrelated diseases, such as diabetes, angiogenesis, stroke, hypertensive arterial hypertrophy, atherosclerosis, restenosis, glomerular nephritis, cancer and so on.
As regards specifically cells of the Central Nervous System (CNS), since late 1950's it has been shown that the mammalian neuronal development is controlled by a family of growth factors, later called neurotrophins, whose most important member appears to be Nerve Growth Factor (NGF). NGF was purified in the early 1970's, and subsequently has been cloned and sequenced. The native peptide consists of three pairs of subunits, of which the active one is a 118 residue beta-subunit. It is now well established that NGF exerts its effects on defined cell populations of neurons within the CNS, where specific receptors have been identified, cloned and sequenced: its mechanism of activity appears related to increased expression of early response genes inside the cells, leading to changes in the genetic program.
The effects of administration of uridine in brain have also been observed, and it has been shown uridine is able to protect against experimental epilexy and other neural pathology. The Applicant, for example, has already obtained a U.S. Pat. No. 4,960,759 for the use of uridine as a drug modulating the effects of dopamine in the Central Nervous System and, more recently, another U.S. Pat. No. 5,190,948 for treatment of the complications of diabetes at the peripheral nervous system level. The effects of administration of uridine however are not limited to Nervous System.
In general, in fact, uridine is a known compound that has been widely studied ever since it was found to be a constituent element of ribonucleic acids. The large variety of pharmacological effects of uridine lies in the fact that this pyrimidinic nucleoside, as well as forming part of the ribonucleic acids, and thus stimulating biosynthesis of proteins inside the cells, also superintends a number of fundamental biochemical processes, such as the reconstitution of glycogen reserves from glucose, detoxification of cells from numerous exogenous components, and biosynthesis of important constituents that are of structural importance for the cell functions, such as glycolipids and glycoproteins.
The effects of administration of uridine, either alone or associated with cytidine (another pyrimidinic nucleoside), have been studied in various organs in experimental animals: for example, it has been seen in a number of studies that on the isolated heart uridine has positive effects on the use of energy reserves, and improves the myocardial functions. At a muscular level, uridine increases glucose pick-up and biosynthesis of glycogen deposits. In the liver it improves hepatic regeneration after experimental intoxication.
Uridine is also one of the most important agents for the recovery of cell functions in mammals, and for this reason its concentration in the plasma is kept at more or less constant levels by means of enzymatic mechanisms, located above all in the liver. When the blood levels are too low (for example, as a result of damage to the liver) the uridine can be formed ex novo using a complex enzymatic system, comprising a transfer of electrons inside the mitochondrions: however, if the mitochondrions are not functioning adequately, as is the case during ageing or as a result of cell intoxication, then the external supply of uridine through the blood stream becomes of fundamental importance, lack of supply resulting in degeneration and subsequent death of the cells involved. Moreover external uridine is quickly and easily taken up through the outside membrane of the cell.
The ability of exogenous uridine to be used rapidly for biosynthesis of cell ribonucleic acids has been used therapeutically for a long time. There are a number of anti-tumour drugs available whose molecular structure is based on that of uridine, so that cells in rapid growth pick them up in large quantities, thus becoming intoxicated by altered chemical functions, which do not allow correct protein biosynthesis. In particular it has been demonstrated experimentally that anti-viral and anti-tumour agents cause important damage at a mitochondrial level (see, for example, Biochemical Pharmacology 38, 1033-1036, 1989, id. 42, 1397-1400, 1992), and are responsible for even fatal side effects, which can be observed in their long-term use (see, for example New England J. Med. 322, 1098-1105, 1990; id. 333, 1146-1148, 1995).
Unfortunately this biological damage is not only caused to the malign cells, but also spreads to healthy cells when these need to produce new proteins, so that use of the above mentioned drugs is strongly limited by their systemic toxicity. To overcome this problem, in order to allow the healthy cells to return to normal activity after intoxication by the anti-tumour agents, in recent years a special therapy has been perfected, which foresees the use of large amounts of uridine immediately after the use of anti-tumour agents such as 5-fluorouracyl, which is considered to be the reference drug for cancer of the colon (see for example Seminars in oncology 19, supp.3, 148-154, 1992).
Likewise important anti-viral agents, such as those currently in use to treat patients suffering from AIDS, are based on structures similar to uridine, in order to "intoxicate" the biosynthesis of viral proteins, or to inhibit the activity of particular enzymes (for example reverse transcriptase). In this case also, however, healthy cells are adversely affected, which theoretically might be prevented by administration of uridine: it has in fact been demonstrated "in vitro" on cell cultures that uridine is capable of abolishing the toxicity of azidodeoxythymidine (AZT) in human cells producing bone marrow (antimicrobial Agents and Chemotherapy 32, 997-1001, 1988) and, alongside pyruvate, that of Dideoxycytidine in PC12 cells (Molecular Pharmacology 44, 702-706, 1993). In spite of these results no drug composition based on uridine is actually in trade.
The reasoning behind the use of uridine, both in the case of the anti-tumour drug 5-fluorouracyl, and in case of the anti-viral drugs AZT and dideoxycytidine, is that there is competition on the part of the cells to pick up the pyrimidinic nucleosides, and even if it is not possible to reach an ideal situation in which the harmful cells pick up the pharmaceutical agents, while the healthy ones use the uridine, however it can be supposed that, by administering high doses of uridine after the drugs, the healthy cells can return to a normal level of activity, "pushing out" by competition the drugs from the sites they occupy in the ribonucleic acids.